Congenital heart disease is frequently due to abnormalities in cardiac development. The goal of this research project is to improve our understanding of the molecular events in cardiogenesis. The plan is focused on understanding the transcriptional regulation and function of the transcription factors dHAND and eHAND. A substantial body of data shows that these transcription factors are essential for cardiac morphogenesis and are important mediators of cardiac segment specification. The proposal has three experimental Aims. Aim 1 examines structure-function relationships of dHAND by asking if the closely related protein eHAND can functionally replace it. If not, a followup experiment will ask if the difference is due to the divergent DNA binding domain of eHAND. Aim 2, we propose to identify proteins that interact with dHAND. This information will help us understand how dHAND regulates formation of the heart. This screen is also likely to uncover novel regulators of heart formation. Aim 3 proposes to determine the regulatory sequences which govern eHAND expression in the developing heart, particularly at the heart tube stage. EHAND is one of the first transcription factors known to be differentially expressed in the developing heart. Defining cis-acting regions which regulate eHAND will be a crucial first step towards defining the signaling pathways and transcription factors that act through these sequences to pattern the heart tube. To achieve these aims, we will use engineered mice to alter dHAND coding sequences and the eHAND promoter. The effect of these mutations will be analyzed in engineered mouse embryos. The mouse system has been chosen because of the similarity of mouse and human heart development, and because the mouse genome is uniquely amenable to genetic manipulation. Under the preceptorship of a successful physician-scientist who is knowledgeable about vertebrate heart development, the applicant expects to gain the skills necessary to make contributions towards understanding heart development, and how abnormalities in this process lead to congenital heart disease. This experience, and the information generated in this plan, will launch the applicant's career as an independent investigator.